Mitochondrial dysfunction is seen not only in late-onset neurodegenerative disease, such as Alzheimer's, Parkinson's, and Huntington's, but with aging in the normal brain as well. Since the frequency of mitochondrial DNA (mtDNA) mutations in the brain climbs hundreds to thousands of fold with age, it is widely thought that such mutations may contribute to cause mitochondrial dysfunction. To experimentally probe their pathophysiology, transgenic mice were constructed that rapidly accumulate specifically mtDNA mutations in cardiomyocytes. These mice reveal that mtDNA mutations - at frequencies commonly seen with age or disease in humans - indeed cause pathology. Characterization of mitochondria from those mice suggests a novel molecular mechanism for the pathogenesis of elevated levels of mtDNA mutations. As mutations rise so do the levels of mutant proteins encoded by the mitochondrial genome. Some of these mutant proteins will misfold. One of the major chaperones catalyzing protein folding in mitochondria is cyclophilin D (CyP-D), a peptidyl-prolyl cis/trans isomerase that also functions to regulate mitochondrial pore transition. Elevated levels of misfolded mitochondrial-encoded proteins are proposed to lead to dysfunction of CyP-D and, in turn, to dysregulation of pore transition. Catastrophic pore transition is known to cause massive disruption of calcium homeostasis in neurons and to signal cell death by apoptosis. To test these hypotheses, we propose to: 1) characterize the structural and functional alteration in CyP-D that occur when the levels of mtDNA mutations rise, 2) determine the basis for the alteration in mitochondrial pore transition that occurs when mutation levels rise, and 3) generate transgenic mice with an accelerated accumulation of mtDNA mutations in the brain to characterize the effect(s) of these mutations on the function of CyP-D and the permeability transition pore in neurons. These studies are broadly significant to understand molecular mechanisms for the pathogenesis of mtDNA mutations. Since such mutations may be an important contributing factor for many adult-onset diseases, these studies may provide insights into novel therapeutic strategies.